The National Cancer Institute projects that more than 1,000,000 cases of skin cancer will be diagnosed in the United States in 2008, including more than 62,000 new cases of melanoma. Although surgical excision of the primary tumor is curative for the majority of squamous and basal cell skin cancers, approximately 10% of primary or recurrent non-melanoma skin cancers present as complex, perineural, or locally invasive tumors that are more difficult to manage by surgery alone. Tumor recurrence near the site of the primary tumor remains a predominant cause of treatment failure. The ultimate goal of this Fast-Track SBIR proposal is to develop a novel bio-engineered tissue therapeutic that will prevent the spread or recurrence of locally-invasive or complex skin cancers through the sustained, expression of a potent immunostimulatory cytokine at the site of a recently resected tumor. Phase I proposes to generate keratinocyte-specific expression vectors and demonstrate production of bioactive cytokine from transformed keratinocytes. The isolation and characterization of a panel of stably-transfected keratinocytes will identify three candidate clones producing high, medium, and low levels of cytokine expression for further evaluation in Phase II. During Phase II, the most promising clones will be evaluated for karyotype, tumorigenicity, and transgene configuration. Using a murine xenograft model, the ability of these candidate clones to suppress the growth of human tumors in vivo will be assessed. The clone that exhibits the greatest tumor growth suppression will be identified as the leading candidate for further preclinical development. A master cell bank will be prepared from this clone according to current Good Manufacturing Practices. This cell bank will be qualified using a panel of tests recommended by the FDA for characterization of cell lines used to produce biological products. Successful completion of the research outlined in this Fast-Track proposal will generate critical pre-clinical safety and efficacy data that will support the filing of an Investigational New Drug (IND) application with the FDA for evaluation of this technology in human clinical trials. PUBLIC HEALTH RELEVANCE: The National Cancer Institute projects that more than 1,000,000 cases of skin cancer are expected to be diagnosed in the United States in 2008, including more than 62,000 new cases of melanoma. Although most cases of skin cancer can be treated surgically, tumor recurrence near the original tumor site is the leading cause of treatment failure. The goal of this project is to develop a living skin substitute that will express a factor known to inhibit the growth of human tumors, in addition to providing factors that promote the healing of skin wounds. This product has the potential to prevent the spread or recurrence of residual tumor cells following surgical removal of primary skin cancers.